PbGA2ox8 induces vascular-related anthocyanin accumulation and contributes to red stripe formation on pear fruit

Fruit with stripes,which are generally longitudinal,can occur naturally,but the bioprocesses underlying this phenomenon are unclear.Previously,we observed an atypical anthocyanin distribution that caused red-striped fruit on the spontaneous pear bud sport“Red Zaosu”(Pyrus bretschneideri Rehd.).In this study,comparative transcriptome analysis of the sport and wild-type“Zaosu”revealed that this atypical anthocyanin accumulation was tightly correlated with abnormal overexpression of the gene-encoding gibberellin(GA)2-beta-dioxygenase 8,PbGA2ox8.Consistently,decreased methylation was also observed in the promoter region of PbGA2ox8 from“Red Zaosu”compared with“Zaosu”.Moreover,the GA levels in“Red Zaosu”seedlings were lower than those in“Zaosu”seedlings,and the application of exogenous GA4 reduced abnormal anthocyanin accumulation in“Red Zaosu”.Transient overexpression of PbGA2ox8 reduced the GA4 level and caused anthocyanin accumulation in pear fruit skin.Moreover,the presence of red stripes indicated anthocyanin accumulation in the hypanthial epidermal layer near vascular branches(VBs)in“Red Zaosu”.Transient overexpression of PbGA2ox8 resulting from vacuum infiltration induced anthocyanin accumulation preferentially in calcium-enriched areas near the vascular bundles in pear leaves.We propose a fruit-striping mechanism,in which the abnormal overexpression of PbGA2ox8 in“Red Zaosu”induces the formation of a longitudinal array of anthocyanin stripes near vascular bundles in fruit.

Fine-mapping and validation of the genomic region underpinning pear red skin colour

Red skin colour is an important target trait in various pear breeding programmes.In this study,the genetic control of red skin colour was investigated in an interspecific population derived using the descendants of the red sport European pear cultivar‘Max Red Bartlett’(MRB)and the red-blushed Chinese pear cultivar‘Huobali’.Approximately 550 seedlings from nine families were phenotyped for red skin over-colour coverage(Ocolcov)and the intensity of red over-colour(Ocolint)on a 0–9 scale,and genotyped using genotyping-by-sequencing.Genome-wide association analyses were conducted using 7500 high-quality single nucleotide polymorphisms(SNPs).Genomic regions on linkage groups(LG)4 and 5 were found to be associated,and the best SNP(S578_25116)on LG4 accounted for~15%of phenotypic variation in Ocolcov and Ocolint.The association of S578_25116 with Ocolcov and Ocolint was successfully validated in a sample of~200 European and Asian pear accessions.The association with red skin at locus S578_25116 was not present in Asian pear accessions,suggesting its close proximity to the MRB’s Cardinal gene.Several putative candidate genes,including MYB transcription factors(PCP027962 and PCP027967),were identified in the quantitative trait locus region on LG4 and await functional validation.

Competition between anthocyanin and kaempferol glycosides biosynthesis affects pollen tube growth and seed set of Malus

Flavonoids play important roles in regulating plant growth and development.In this study,three kaempferol 3-O-glycosides were identi fi ed and mainly accumulated in fl owers but not in leaves or fruits of Malus.In Malus,fl ower petal color is normally white,but some genotypes have red fl owers containing anthocyanin.Anthocyanin biosynthesis appears to be in competition with kaempferol 3-O-glycosides production and controlled by the biosynthetic genes.The white fl ower Malus genotypes had better-developed seeds than the red fl ower genotypes.In fl owers,the overexpression of MYB10 in Malus domestica enhanced the accumulation of anthocyanin,but decreased that of kaempferol 3-O-glycosides.After pollination the transgenic plants showed slower pollen tube growth and fewer developed seeds.Exogenous application ofdifferent fl avonoid compounds suggested that kaempferol 3-O-glycosides,especially kaempferol 3-O-rhamnoside,regulated pollen tube growth and seed set rather than cyanidin or quercetin 3-O-glycosides.It was found that kaempferol 3-O-rhamnoside might regulate pollen tube growth through effects on auxin,the Rho of plants(ROP)GTPases,calcium and the phosphoinositides signaling pathway.With the inhibition of auxin transport,the transcription levels of Heat Shock Proteins(HSPs)and ROP GTPases were downregulated while the levels were not changed or even enhanced when blocking calcium signaling,suggesting that HSPs and ROP GTPases were downstream of auxin signaling,but upstream of calcium signaling.In summary,kaempferol glycoside concentrations in pistils correlated with auxin transport,the transcription of HSPs and ROP GTPases,and calcium signaling in pollen tubes,culminating in changes to pollen tube growth and seed set.

Extreme-phenotype GWAS unravels a complex nexus between apple(Malus domestica)red-flesh colour and internal flesh browning

The genetic link between apple red flesh(RF)coloration and the internal flesh browning disorder(FBD)is a major challenge when breeding high fruit quality RF apple cultivars.A genome-wide association study(GWAS)was conducted in a population of about 900 red-leaved seedlings to identify genomic regions and putative candidate genes using whole genome sequencing of the pools of extreme phenotypes(XP)for the RF colour coverage(using the weighted cortex index(WCI))and FBD.This study identified novel genomic regions contributing to WCI and FBD variation in the red-leaved seedlings.The FBD-associated regions were enriched for genes regulating senescence,heat shock proteins,cytochrome P450,ascorbate metabolism and pectin methyl esterases.Although there were no significant regions in common for WCI and FBD,there were several genes(e.g.MYB85,MYB66,ethylene insensitive 3,DNAJ heat shock protein,WRKY7,and NAC42)enriched commonly between the genomic regions associated with these traits,potentially underpinning the genetic connection between WCI and FBD.Some of the differentially expressed genes between the R6:MdMYB10 and white-fleshed‘control’apples resided within the GWAS hotspot for WCI(e.g.chalcone synthase,UDP-Glycosyl transferase)and FBD(e.g.Rho GTPase activating protein,lipoxygenase 1,phytoene synthase)–validating the XP-GWAS findings.Paralogs of several genes resided in the trait-associated genomic regions,suggesting that whole genome duplication plays an important role in the regulation of these traits.Adverse genetic correlations between WCI and sensory traits were observed,and strategies to develop FBD-free high fruit quality RF cultivars are discussed.

Unraveling a genetic roadmap for improved taste in the domesticated apple

Although taste is an important aspect of fruit quality, an understanding of its genetic control remains elusive in apple and other fruit crops. In this study, we conducted genomic sequence analysis of 497 Malus accessions and revealed erosion of genetic diversity caused by apple breeding and possible independent domestication events of dessert and cider apples. Signatures of selection for fruit acidity and size, but not for fruit sugar content, were detected during the processes of both domestication and improvement. Furthermore, we found that single mutations in major genes affecting fruit taste, including Ma1, MdTDT, and MdSOT2, dramatically decrease malate, citrate, and sorbitol accumulation, respectively, and correspond to important domestication events. Interestingly, Ma1 was identified to have pleiotropic effects on both organic acid content and sugar:acid ratio, suggesting that it plays a vital role in determining fruit taste. Fruit taste is unlikely to have been negatively affected by linkage drag associated with selection for larger fruit that resulted from the pyramiding of multiple genes with minor effects on fruit size. Collectively, our study provides new insights into the genetic basis of fruit quality and its evolutionary roadmap during apple domestication, pinpointing several candidate genes for genetic manipulation of fruit taste in apple.